Abstract:

The present invention relates to aerosols containing magnetic particles,
wherein the aerosols comprise magnetic particles and a pharmaceutical
active agent. The invention furthermore relates to the use of such
aerosols containing magnetic particles for directed magnetic field-guided
transfer of the active agents contained therein in aerosol therapy.

Claims:

1. Aerosol containing magnetic particles, wherein the aerosol contains
magnetic particles having a diameter of at least 5 nm and at most 800 nm
and at least one pharmaceutical active agent.

2. The aerosol containing magnetic particles according to claim 1, wherein
the magnetic particles have a diameter of at least 50 nm and at most 750
nm, preferably of at least 100 nm and at most 700 nm, more preferably of
at least 150 nm and at most 600 nm, still more preferably of at least 200
nm and at most 500 nm, particularly preferably of at least 250 nm and at
most 450 nm, most preferably of at least 300 nm and at most 400 nm.

3. The aerosol containing magnetic particles according to claim 1, wherein
the magnetic particles and the pharmaceutical active agent are contained
in a solvent.

4. The aerosol containing magnetic particles according to claim 3, wherein
the solvent is an inorganic or organic solvent.

5. The aerosol containing magnetic particles according to claim 3, wherein
the solvent is selected from the group consisting of ethanol, water and
glycerine and mixtures thereof.

6. The aerosol containing magnetic particles according to claim 1, wherein
the pharmaceutical active agent is coupled to the magnetic particles.

7. The aerosol containing magnetic particles according to claim 1, wherein
the pharmaceutical active agent is selected from the group consisting of
nucleic acids, peptides, proteins, cytostatics, broncholytics,
antibiotics, antidiabetics and immunomodulators.

8. The aerosol containing magnetic particles according to claim 1, wherein
the pharmaceutical active agent is contained in a vector, liposome,
hollow colloid or nanoparticle.

10. The aerosol containing magnetic particles according to claim 1,
wherein the magnetic particles consist of metals, or contain these,
selected from the group consisting of iron, cobalt or nickel, magnetic
iron oxides or hydroxides, such as Fe3O4,
gamma-Fe2O3, double oxides or hydroxides of di- or trivalent
iron ions with other di- or trivalent metal ions, such as Co2+,
Mn2+, Cu2+, Ni2+, Cr3+, Gd3+, Dy3+ or
Sm3+, and any mixtures thereof.

11. The aerosol containing magnetic particles according to claim 1,
wherein the magnetic particles consist of paramagnetic or
superparamagnetic material or contain this.

13. The pharmaceutical composition according to claim 12, additionally
comprising at lease one (further) solvent, at least one complexing agent
and/or at least one pharmaceutically acceptable acid.

14. A method for prophylaxis and/or therapy of diseases of the respiratory
tract and/or lungs, inflammatory and/or obstructive diseases of the
respiratory tract and/or lungs selected from melanomas or malignant
melanomas of the respiratory tract, the lung, lung cancer, lung tumours,
lung carcinomas, small cell lung carcinomas, throat cancer, bronchial
carcinomas, larynx cancer, head/neck tumours, tongue cancer, sarcomas and
blastomas in the region or vicinity of the respiratory tract, the lung,
as well as asthma, COPD (chronic obstructive pulmonary disease), lung
emphysema, chronic bronchitis, pneumonia and hereditary diseases,
mucoviscidosis, human surfactant protein B deficiency and
α1-antitrypsin deficiency, and for therapy after a lung
transplant and for pulmonary vaccination and for anti-infective therapy
of the lung comprising administering an effective amount of the aerosol
containing magnetic particles of claim 1 to a subject in need thereof.

15. (canceled)

16. The method according to claim 14, wherein the aerosol containing
magnetic particles is deposited by a magnetic field onto the surface of
the region of the respiratory tract and/or lung to be treated.

17. The method according to claim 16, wherein the magnetic field has a
field strength of at least 100 mT (millitesla), at least 200 mT, at least
500 mT or at least 1 T (tesla).

18. The method according to claim 16, wherein the magnetic field has a
magnetic field gradient of greater than 1 T/m or greater than 10 T/m.

19. The method according to claim 14, wherein the magnetic field is a
pulsating, an oscillating or pulsating-oscillating magnetic field.

20. The method according to claim 14, wherein the magnetic field is
matched dynamically to the breathing of the patient and is active only
during the resting pauses between in- and exhalation of ex- and
inhalation.

21. A kit comprising an aerosol containing magnetic particles according to
claim 1, and external which generates a magnetic field and a nebulizer.

[0002]Aerosols are solid or liquid suspended particles in gases (in
particular air) having a diameter of from about 0.0001 μm to about 100
μm, it being possible for the composition and form of aerosols to vary
very greatly. If solid particles are present in the aerosol, such
aerosols are typically called smokes or dusts, whereas if liquid
particles are present in the aerosol, these aerosols are typically called
mists. In addition, mixed forms of these aerosols can also occur, i.e.
aerosols with solid and liquid suspended particles. In recent decades
numerous synthetic aerosols have been prepared for a wide industrial and
commercial field of use. Such synthetic aerosols can be prepared by
conventional dispersion and condensation processes and are as a rule used
in spray cans in combination with a liquefied compressed gas as the
propellant gas. Depending on the nature of the particles contained
therein, they are used, for example, as hair and body care compositions,
deodorants, perfumes, odour improvers, disinfection and pest control
compositions, flooring, glass and furniture care compositions, lacquers
and paints, automobile care compositions etc. Aerosols are also used in
particular, with and without propellant gas, in the field of medicine, in
so-called aerosol therapy for treatment of various diseases of the
respiratory tract and/or lungs. Pharmaceutical active agents, e.g.
salbutamol, formoterol, ipatropium bromide, budesonide, fenoterol,
terbutaline, tiotropium bromide, salmeterol, beclometasone, fluticasone,
mometasone, tobramycin, theophylline, dornase
α,α1-antitrypsin, interferon-β, insulin, calcitonin
or growth hormones, can be administered via the lungs by means of the
medicinal aerosols mentioned last. The present invention and following
description relate to such medicinal aerosols.

[0003]The smallest (pharmaceutically active) particles in medicinal
aerosols are e.g. nucleic acids, peptides or proteins, while the largest
particles are e.g. mist particles. The aerosols often comprise mixtures
of particles of different particles sizes and therefore embody a
polydisperse size distribution. When considering the size distribution of
aerosols, it is generally important whether the number, the surface or
the weight of the particles is under consideration (it being noted in
this connection that, for example, a particle of diameter 10 μm
corresponds to the weight of 1,000 particles of diameter 1 μm). The
size distribution spectrum is generally specified by a parameter called
the mass median aerodynamic diameter (MMAD), as a rule 50% of the aerosol
mass being larger and 50% smaller than the MMAD. In this connection, it
is to be noted that for biological systems in particular, the mass
aerodynamic distribution of the aerosol spectrum is as a rule used, such
as is described, for example, by Kohler, D. & Fischer, F. in Theorie und
Praxis der Inhalationstherapie [Theory and Practice of Inhalation
therapy] (Arcis Verlag GmbH, Munich, 2000).

[0004]Medicinal aerosols in aerosol therapy are typically inhaled orally
or nasally by the patient to be treated. During or after inhalation of
the particles into the lung a certain proportion of the particles escapes
from the flow line of the particles formed by the in- or exhalation and
thereby comes into contact with the moist surface of the air cavities,
e.g. the throat, nasal or pharyngeal cavity, the trachea or the lung
tissue. This phenomenon is in general called particle deposition or
deposition and is subject in particular to the following three physical
mechanisms: [0005]impaction, [0006]sedimentation and [0007]diffusion

[0008]In impaction, the aerosol particles up to a certain diameter follow
the flow line of the inhaled aerosol. Above a diameter of from 2 to 3
μm, the inertia of the aerosol becomes relevant, which means that the
aerosol particles have the tendency to fly straight on when the flow line
of the aerosol changes direction, and to be deposited on the surface.
Such changes in direction of the flow line of the aerosol take place in
particular due to the physiological shape of the respiratory tracts, e.g.
the oropharynx, the branchings of the respiratory tract to the left and
right lung lobe and/or the branchings in the region of the alveoli. The
deposition probability (DE) for impaction is proportional to the square
of the diameter (d) and the aerosol flow (V):

DE˜d2V

[0009]Impaction is an important deposition mechanism for aerosols in the
size range above a particle diameter of 3 μm. Deposition by impaction
is therefore to be found everywhere where high aerosol speeds and marked
changes in direction occur, as is mostly the case in the wide respiratory
tract and the oropharynx. Larger particles above 10 μm are deposited
above all to the extent of more than 90% at the first marked change in
direction, i.e. the oropharynx (see e.g. Kohler, D. & Fischer (2000,
supra); Schulz, H. & Muhle, H. 323-345 (Academic Press, 2000)).

[0010]Sedimentation is a deposition mechanism which is to be attributed to
the gravity of the aerosol particles. Here also, deposition depends on
the particle size and takes place in particular at a particle diameter
above from 0.5 to 1 μm. The settling speed (vs), which is the
determining factor here, can be described approximately by the following
equation:

[0012]In contrast to impaction and sedimentation, in diffusion, particles
having a diameter of smaller than 1 μm on the one hand still follow
the gas stream, i.e. the flow line, and on the other hand more and more
resemble molecules which are subjected to molecular Brownian motion. Due
to diffusion, in the vicinity of the wall these particles can therefore
leave their initial flow line and be deposited on the wall. The average
advancement of a particle (Δ) during diffusion is calculated as
follows:

Δ = 2 k T C t π η d ##EQU00002##

[0013]The average advancement of a particle (Δ) during diffusion
therefore depends on the particle diameter (d), the viscosity of the gas
(η), the time (t) and the temperature (T in ° K.). The
constants of the equation are: C=Cunningham's slip correction and
k=Boltzmann constant. The diffusion distance somewhat resembles a
bell-shaped frequency distribution around the starting value, the average
advancement being proportional to the root of the variable (see e.g.
Kohler, D. & Fischer (2000, supra)).

[0014]Table 1 gives a comparative overview of the distances covered with
respect to sedimentation and diffusion. It can be seen that small
particles can cover considerable distances via diffusion.

[0015]For therapeutic use of aerosols, the deposition mechanism is of
decisive importance for the choice of the particle size of various active
agents for therapy and/or prophylaxis of diverse diseases of the
respiratory tract and/or lungs. In addition, however, the matter of the
regions in which the inhaled particles are to be deposited in the lung is
of considerable importance. Detailed studies in the prior art have shown
that under certain circumstances (e.g. the particle size described above
or the breathing technique, i.e. the nature and manner of breathing)
deposition takes place preferentially in certain compartments of the
bronchial tree or alveolar region. For effective and gentle therapy it is
desirable for the aerosols and the pharmaceutical active agents
transported by these aerosols to be administered in a targeted manner to
only defined, diseased regions of the lungs. On the one hand, the dose of
the pharmaceutical active agent to be administered can be reduced by this
means, and on the other hand undesirable side effects on the surrounding
healthy tissue can be reduced or avoided.

[0016]To achieve this aim, the problem emerges that the natural or
"normal" spreading of the inhaled aerosols (e.g. in the bronchial tree or
alveolar region) would have to be influenced if administration to other
regions is desired in order to be able to achieve a directed spreading
into such defined regions of the lung. In this connection, various
mechanisms for directed supply of aerosol into the lung have been
proposed in the prior art, such as, for example, described by Ernst, N.
et al. (Interaction of liposomal and polycationic transfection complexes
with pulmonary surfactant. J Gene Med 1, 331-40. (1999)) and Rosenecker,
J. et al. (Interaction of bronchoalveolar lavage fluid with polyplexes
and lipoplexes: analysing the role of proteins and glycoproteins. J Gene
Med 5, 49-60. (2003)). Thus, for example, changes in the particle size in
combination with various breathing techniques, e.g. long holding of
breath or also an extremely slow inhalation of aerosols, have been
proposed. The individual respiratory tract geometry of the patient has
likewise been taken into account. For certain active agents, e.g. for a
DNA transfer in the context of gene therapy, the use of viral or
liposomal vectors in epithelial cells of the respiratory tract for
treatment of mucoviscidosis has furthermore been described (Rudolph, C.
et al. Nonviral gene delivery to the lung with copolymer-protected and
transferrin-modified polyethylenimine. Biochim Biophys Acta 1573, 75-83.
(2002)).

[0017]Ally et al. (Journal of Magnetism and Magnetic Materials 293,
442-449 (2005)), describe hypothetically the possibility of a
magnet-guided transportation of aerosol particles. For this, guiding of
chemotherapeutic active agents with the aid of a magnetic field into
regions of the lung affected by lung cancer is proposed. However, the
studies are based only on an in vitro model in which solid carbonyl-iron
particles having a diameter of from 1 to 3 μm are used in air. The
particle speed of v=0.34 m/s used here in combination with the magnetic
field strength of 36 mT for the stated particle size of from 1 to 3 μm
is coordinated to the in vitro studies described, however, and cannot be
applied to the deviating in vivo conditions. Studies of the present
invention have shown that under the conditions chosen by Ally et al.
(2005, supra), only a very inadequate transportation of particles takes
place in vivo. in vivo conditions which do not arise in an in vitro
system, such as impeding of the transportation of aerosol by
physiological conditions, enzymes, mucous membranes, structure and
construction of the respiratory tract etc., moreover are not taken into
account by Ally et al. (2005, supra). In other words, effective
transportation of particles in vivo, such as is required in aerosol
therapy, cannot be achieved with the parameters stated in Ally et al.
(2005, supra).

[0018]Summarizing, it is to be said that none of the procedures described
in the prior art has led to targeted transportation and directed
deposition of aerosol particles and therefore of pharmaceutical active
agents in defined regions of the lung being ensured. This has the
disadvantage that increased amounts of pharmaceutical active agents must
be administered in order to achieve the intended action in the diseased
tissue to be treated in the lung, which as a result leads to increased
active agent costs and consequently also to increased therapy costs. A
further serious disadvantage is that the aerosols loaded with active
agent not only are deposited in diseased regions of the respiratory tract
or lung, but are also deposited in regions which are not affected by the
disease, i.e. in healthy tissue. This deposition pattern is a
disadvantage, since undesirable side effects may occur on non-diseased
tissue of the respiratory tract or lungs due to contact with or uptake of
the active agent administered. These considerations illustrate that novel
methods which render possible a directed local deposition of aerosols in
the respiratory tract and in particular in the lung are required.

[0019]The object of the present invention is to provide a system by which
a directed in vivo transportation of aerosols into defined regions of the
respiratory tract and of the lung is ensured.

[0020]This object is achieved by the subject matter of claim 1 of the
present invention. Advantageous embodiments of the invention are
described in the further claims.

[0021]The present invention relates in first subject matter to an aerosol
containing magnetic particles, wherein the aerosol contains magnetic
particles having a diameter of at least 5 nm and at most 800 nm and at
least one pharmaceutical active agent. Preferably, the magnetic particles
contained therein have a diameter of at least 50 nm and at most 750 nm,
further preferably of at least 100 nm and at most 700 nm, more preferably
of at least 150 nm and at most 600 nm, still more preferably of at least
200 nm and at most 500 nm, particularly preferably of at least 250 nm and
at most 450 nm, most preferably of at least 300 nm and at most 400 nm.

[0022]The invention is based on studies with which it was possible to
demonstrate for the first time that an inhaled aerosol according to the
invention containing magnetic particles which contains a pharmaceutical
active agent can be transported in a directed, i.e. targeted, manner into
defined regions of the lung. The transportation of this aerosol according
to the invention containing magnetic particles into defined regions of
the lung takes place via an externally applied magnetic field which
causes deposition of the magnetic particles and consequently also of the
aerosol on the surface of the desired region of the lung. The present
invention therefore provides an effective aerosol which can be used in
vivo for directed transportation of active agent in aerosol therapy. In
this connection, the term "in vivo" means any use of the aerosol
according to the invention containing magnetic particles on the body of a
living multi-cell organism, preferably a mammal, more preferably a human.
In contrast to this, in this connection "in vitro" means any use of the
aerosol according to the invention containing magnetic particles outside
such a body or organism.

[0023]The magnetic particles contained in the aerosol according to the
invention containing magnetic particles can consist of various metals and
oxides or hydroxides thereof or contain these. Magnetic particles which
are suitable according to the invention are described, for example, in
the international patent application WO 02/000870, the disclosure content
of which in this respect is subject matter of the present invention. The
term "magnetic particles" means magnetically reacting solid phases. These
solid phases are typically particles or aggregates thereof having a
diameter in the nano- to micrometer range of not larger than 800 nm, and
conventionally contain one or more metals or oxides or hydroxides thereof
which react to the magnetic force of a magnetic field and are preferably
attracted or accelerated in a defined direction by the source of the
magnetic field. Temporarily magnetic particles, for example of
ferrimagnetic or, preferably, ferromagnetic materials, are likewise
included. Particles of paramagnetic or superparamagnetic material are
furthermore included in the present invention. Suitable materials of the
magnetic particles include, for example, iron, cobalt or nickel, magnetic
iron oxides or hydroxides, such as Fe3O4,
gamma-Fe2O3, or double oxides or hydroxides of di- or trivalent
iron ions with other di- or trivalent metal ions, e.g. Co2+,
Mn2+, Cu2+, Ni2+, Cr3+, Gd3+, Dy3+ or
Sm3+, and any mixtures of such oxides or hydroxides. Preparation
processes for magnetic particles are described e.g. in Schwertmann U. and
Cornell R. M., Iron Oxides in the Laboratory, VCH Weinheim 1991, in WO
02/000870 and in DE 196 24 426.

[0024]The magnetic particles contained in the aerosol according to the
invention containing magnetic particles are typically synthetic magnetic
particles, i.e. are not obtainable from a biological source (a living
organism). Preferably, the magnetic particles or aggregates thereof
induce no systemic toxic side effects in the organism to which they are
administered. According to one embodiment of the invention, the magnetic
particles contained in the aerosol according to the invention containing
magnetic particles are coupled to any vectors, liposomes, hollow colloids
or nanoparticles described herein or to the pharmaceutical active
agent(s) itself/themselves. The magnetic particles of the present
invention contained in the aerosol according to the invention containing
magnetic particles can be present in non-coated or coated form. If the
magnetic particles are present in coated form, the coating is preferably
selected from positively or negatively charged electrolytes, such as
phosphates, citrates or amines, with silanes, fatty acids or polymers,
e.g. polysaccharides, proteins or natural or synthetic polymers. Such a
coating of the magnetic particles contained in the aerosol according to
the invention containing magnetic particles serves, for example, for
reduction of any toxicity of the magnetic particles, for coupling of the
pharmaceutical active agent(s) to the magnetic particles, for
improving/increasing the passage (of the active agent, optionally
together with the magnetic particle(s)) through membranes, etc. Examples
of such coatings are described, inter alia, in U.S. Pat. No. 4,554,088,
U.S. Pat. No. 4,554,089, U.S. Pat. No. 4,208,294, U.S. Pat. No. 4,101,435
and DE 196 24 426, the disclosure content of which in this respect is
included in full in the present invention. These coatings and the
compounds used for them can have reactive functional groups as described
in the following. However, these reactive functional groups can also be
introduced as required by conventional chemical modifications after the
coating operation. Such functional groups can have cation exchange
properties, such as, for example, xanthate, xanthide, dicarboxyl,
carboxymethyl, sulfonate, sulfate, triacetate, phosphonate, phosphate,
citrate, tartrate, carboxylate or lactate groups of natural or synthetic
polymers, such as polysaccharides, polyvinyl alcohol (PVA),
polyvinylpyrrolidone (PVP) or polyethylene glycol (PEG). These functional
groups can be incorporated e.g. into the natural or synthetic polymers
described above before or after the coating of the magnetic particles.

[0025]As already described above, the directed transportation of the
aerosol according to the invention containing magnetic particles, i.e.
the transportation of the magnetic particles and active agent(s), takes
place via an externally applied magnetic field, with which the inhaled
magnetic particles and the pharmaceutical active agent(s) are guided into
the defined regions of the respiratory tract, preferably the lung. A
"magnetic field" which is suitable in the context of the invention
relates to a magnetic field which is generated by a magnet as the source
and, depending on the form and field strength, is capable of attracting
the magnetic particles according to the invention together with active
agent(s) against other physical phenomena acting on these. Such other
phenomena can be, for example, the diffusion, sedimentation and/or
impaction processes described above. Suitable magnetic field for the in
vivo uses according to the invention should preferably generate a field
strength of at least 100 mT (millitesla), preferably of at least 200 mT,
likewise preferably of at least 500 mT, furthermore preferably of at
least 1 T (tesla) and more. The magnetic field gradient generated by the
magnetic fields should preferably be greater than 1 T/m, more preferably
greater than 10 T/m. A "magnet" in the context of the present invention
which generates such magnetic fields described above can be any magnet
suitable for this. For example, permanent magnets or electromagnets
(operated by electric current) can be employed in the context of the
present invention. The intensity (the field strength of the magnet) is
typically controlled via suitable measurement and control instruments
connected to the magnet.

[0026]According to a preferred embodiment, the externally applied magnetic
field is permanently present, i.e. after administration the aerosol
according to the invention containing magnetic particles can be deflected
at any point in time out of the flow line track caused by the in- and
exhalation and deposited on the surface of the respiratory tract, as long
as the magnetic particles contained therein arrive in a region of the
magnetic field strength which is sufficient for this. Such a permanent
magnetic field can be generated by any of the magnets described here, and
preferably has the general properties of a magnetic field as described
above. Preferably, the permanent magnetic field is active for at least
the period of the treatment, i.e. of the administration or inhalation of
the aerosol according to the invention containing magnetic particles.

[0027]According to another preferred embodiment, the externally applied
magnetic field is not permanently present, and is active preferably only
for a part of the period of treatment, i.e. a part of the period of
administration or inhalation of the aerosol according to the invention
containing magnetic particles. More preferably, the magnetic field is
active only during the period of the resting phases between inhalation
and exhalation or between exhalation and inhalation. Such a non-permanent
activation of the magnetic field preferably ensures deposition of the
aerosol according to the invention containing magnetic particles on the
surface during these resting phases and therefore renders possible a
uniform distribution of the aerosol according to the invention containing
magnetic particles. According to a particularly preferred embodiment, the
control of the magnetic field can take place dynamically in coordination
with the breathing of the patient as a function of the breathing rhythm
of the patient, so that during the in- and exhalation by the patient no
magnetic field is applied in the region to be treated, but in the resting
phases a magnetic field is applied there and only then does a deposition
of the aerosol according to the invention containing magnetic particles
on the surface of the respiratory tract take place. Preferably, for this
the air above the chosen surface of the respiratory tract is saturated by
the aerosol according to the invention containing magnetic particles such
that within several activations of the magnetic field(s) by the magnet(s)
a significant deposition of the transported active agent or of the
aerosol according to the invention containing magnetic particles on the
chosen surface of the respiratory tract is rendered possible. Control of
the magnet can be rendered possible, for example, by an electric circuit
which triggers a signal in the nebulizer or inhaler at the start and end
of the inhalation or exhalation of the patient, by which in turn the
magnetic field of a magnet as described here is switched on or off. The
switching on or off of the magnetic field can take place e.g. by
mechanical removal or turning away of the poles of the magnet in the case
of permanent magnets, and likewise mechanically in the case of
electromagnets, but preferably e.g. by switching on or off of the
electric current required for generation of the magnetic field.

[0028]According to a further preferred embodiment, the externally applied
magnetic field is a pulsating magnetic field. A "pulsating magnetic
field" in the present case means in particular that the field strength of
the magnetic field decreases or increases in the region to be treated,
conventionally periodically or virtually periodically. In this context,
the maximum desired field strength is reached in the maximum of the
pulse, while in the minimum of the pulse preferably a lowest possible
field strength, more preferably a field strength less than 20%, still
more preferably a field strength less than 10% of the previously applied
field strength, and still further preferably no field strength is
applied. Particularly preferably, the pulsating magnetic field is
coordinated dynamically with the breathing of the patient as a function
of the breathing rhythm of the patient such that the maximum of the pulse
lies in a resting phase between inhalation and exhalation or between
exhalation and inhalation, while the minimum of the pulse lies during the
inhalation or exhalation. In this context, the pulsating magnetic field
can have the "profile" of a rectangular pulse, a sinusoidal pulse etc.,
or approximations of these profiles. The pulse can be effected here, as
above, by switching on or off of the magnetic field, e.g. by mechanical
removal or turning away of the poles of the magnet in the case of
permanent magnets, and likewise mechanically in the case of
electromagnets, but preferably e.g. by switching on or off of the
electric current required for generation of the magnetic field. The
pulsating magnetic field can furthermore be generated with direct current
or with alternating current if an electromagnet is used. If an
electromagnet is operated with direct current, a magnetic field which
does not change its direction (+/- poling) is preferably generated. In
this context, the magnetic field formed can be adjusted by a person
skilled in the art as required, according to the poling (+/- or -/+
poling).

[0029]According to a further preferred embodiment, the externally applied
magnetic field is an oscillating magnetic field. In the context of the
present invention, the term "oscillating magnetic field" is to be
understood as meaning a magnetic field which periodically changes its
direction (+/- poling). Such an oscillating magnetic field is typically
generated by using an electromagnet and operating the electromagnet with
alternating current. A change in the direction of the magnetic field
(i.e. change in the +/- poling) effected by an oscillating magnetic field
can preferably exert kinetic energy on the magnetic particles, under the
influence of which the transportation of the aerosol according to the
invention containing magnetic particles and/or, for example, the release
of the pharmaceutical active agent coupled to the magnetic particles in
the aerosol according to the invention containing magnetic particles is
promoted, effected or accelerated. An oscillating magnetic field can be
permanently present, or, as described above, can be matched to the
breathing rhythm of the patient such that during the in- and exhalation
by the patient, no oscillating magnetic field is applied in the region to
be treated, but an oscillating magnetic field is applied there in the
resting phases.

[0030]All the embodiments described above can also be combined with one
another in a suitable manner. Thus, for example, a pulsating magnetic
field can be operated in oscillation (pulsating oscillating magnetic
field), i.e. the magnetic field has a characteristic pulse (e.g.
rectangular or sinusoidal pulse) which, for example, oscillates in its
maximum or has an oscillating course from the maximum to the minimum
which constantly decreases in field strength etc.

[0031]The directed transportation of the aerosol according to the
invention containing magnetic particles, i.e. the transportation of the
magnetic particles and active agent(s), takes place by means of an
externally arranged magnet, i.e. outside the organism to be treated
(mammal, preferably human). The directed transportation into defined
regions of the respiratory tract, e.g. the lung, takes place after
inhalation of the aerosol according to the invention containing magnetic
particles by the organism to be treated (mammal, preferably human)
preferably via corresponding change(s) in position of the external
magnets to the defined regions. Accordingly, it is advantageous if the
magnet is freely movable. Freely movable means, for example, that the
magnet can be led/moved manually. However, it is also possible and
advantageous for the magnet to be attached movably to a device, e.g. a
frame, on which its position can be changed, in particular can be
swivelled, adjusted in height and locked, manually, electronically or by
computer control. In this connection, a further possibility for
positioning the external magnet(s) is arrangement of several magnets,
e.g. by an arrangement of permanent magnets or electromagnets, in a
row/in rows or as a bow or as a "sandwich" construction, these magnets
preferably covering the region to be treated. The magnets can be
activated either successively or simultaneously. By such an arrangement
of several magnets e.g. a larger region can be selected for deposition of
the aerosol according to the invention containing magnetic particles and
therefore of the region to be treated.

[0032]The speed of the magnetic particles and pharmaceutical active agents
of the aerosol according to the invention containing magnetic particles
in the lung required for an effective directed transportation depends on
several factors, for example on the regions of the lung into which the
inhaled aerosol is to be transported, on the diameter of the magnetic
particles, on the size of the pharmaceutical active agent component(s),
the individual respiratory tract geometry of the organism treated etc.
The speed can therefore be influenced, for example, by the diameter of
the magnetic particles, the size of the active agent, the breathing
technique, e.g. fast or slow inhalation, deep or shallow breathing,
holding of the inhaled breath, and the field strength applied to the
external magnet(s), or a magnetic field applied in oscillation and/or
pulsation, as described above. For example, a cause of the increase of a
deposition of particles with increasing respiratory minute volume is the
resultant increasing inspiration flow (flow on inhalation). An
end-inspiratory (taking place at the end of the exhalation)
breath-holding time also leads to an increase in the deposition of
aerosol. In the speed (of the aerosol according to the invention
containing magnetic particles and of the magnetic particles and
pharmaceutical active agents according to the invention contained
therein), a distinction is to be made between the speed with which the
aerosol according to the invention containing magnetic particles is
administered, i.e. nebulized (and inhaled), and the speed with which the
aerosol moves into the respiratory tract after administration. The speed
with which the aerosol moves into the respiratory tract after
administration is determined in particular by physiological factors, such
as, for example, the respiratory tract geometry, e.g. the diameter of the
respiratory tract of the patient to be treated, and is e.g. typically
about 4.7 m/s at the second branching of the respiratory tract. The speed
with which the aerosol and therefore the magnetic particles and
pharmaceutical active agents of the invention is/are administered should
advantageously be at least 3 m/s, preferably at least 5 m/s, more
preferably at least 8 m/s, still more preferably at least 10 m/s, in
order to ensure effective transportation of the aerosol according to the
invention containing magnetic particles in vivo. The person skilled in
the art will be able to define the particular suitable speed for
administration of the aerosol according to the invention containing
magnetic particles taking into account the abovementioned factors.

[0034]The pharmaceutical active agent contained in the aerosol according
to the invention containing magnetic particles likewise includes any
cytostatics which are suitable for treatment of diseases of the
respiratory tract and/or lungs. In the present case, "cytostatics" are to
be understood as meaning above all those compounds which have a toxic
action on endogenous cells in a general manner, and inhibit cell growth
in this way. Chemotherapeutics against lung cancer diseases are to be
mentioned here in particular. A distinction is made between cytostatics
of various groups, depending on their action mechanism. By way of
example, the following e.g. are also included here: [0035]Alkylating
and crosslinking cytostatics which damage DNA. Examples of these are
cyclophosphamide, N-nitroso compounds, such as carmustine, ethyleneimine
(aziridine) derivatives, such as thiotepa, methanesulfates, such as
busulfan, platinum complexes, such as cisplatin, procarbazine and others;
[0036]Cytostatic antibiotics, for example anthracyclines, such as
daunorubicin, doxorubicin, bleomycin and mitomycins. The latter
intercalate in DNA and inhibit topoisomerases; [0037]Antimetabolites,
which displace natural metabolism units. Examples are folic acid
antagonists, such as methotrexate, nucleoside analogues, such as
mercaptopurine, fluorouracil and others; and [0038]Hormones and hormone
antagonists. These are employed in particular on tumours of
hormone-dependent growth. Examples are (anti)oestrogens, such as
formestane, gestagens and antiandrogens.

[0039]The pharmaceutical active agents of the aerosols of the present
invention containing magnetic particles can be present in a preformulated
manner, for example packed in suitable agents for transportation of
pharmaceutical active agents, so-called "drug delivery" systems, for
example in nanoparticles, vectors, preferably gene transfer vectors,
viral or non-viral vectors, poly- or lipoplex vectors, liposomes or in a
hollow colloid (i.e. hollow beads of colloidal dimensions). However, they
can also be naked nucleic acids, in particular naked DNA. Suitable
vectors, liposomes, hollow colloids or nanoparticles and processes for
the introduction of substances, such as the pharmaceutical active agents
according to the invention, into such vectors, liposomes, hollow colloids
or nanoparticles are generally well-known in the prior art and are
described, for example, in Cryan S-A. (Carrier-based Strategies for
Targeting Protein and Peptide Drugs to the Lungs, AAPS Journal. 2005;
07(01): E20-E41) and in Sambrook et al. Molecular Cloning A Laboratory
Manual, Cold Spring Harbor Laboratory (1989) NY. Gene transfer vectors
which can be used are, preferably, polyethylenimines or cationic lipids,
such as e.g. DOTAP. Liposomes can preferably be used for packing of
cytostatics (e.g. dilauroylphosphatidylcholines); a detailed description
is given, for example, in Koshkina, N. V. et al. (Paclitaxel liposome
aerosol treatment induces inhibition of pulmonary metastases in murine
renal carcinoma model. Clinical Cancer Research 7, 3258-3262 (2001)).
Proteins as pharmaceutical active agents can preferably be packed into
biocompatible poly-lactic/glycollic acid polymers (PLGA) by means of
supercritical liquids, emulsion processes and spray drying.

[0040]The pharmaceutical active agents of the aerosol according to the
invention containing magnetic particles can likewise be coated with a
magnetic layer. The material of such a magnetic layer preferably consists
of or comprises one of the materials described above for magnetic
particles. Processes for such coatings are known in the prior art and
belong to general technical knowledge.

[0041]The pharmaceutical active agents of the aerosol of the present
invention containing magnetic particles can be present in a form coupled
to or adsorbed on the magnetic particles or not coupled to or adsorbed on
these. In the case of coupling or adsorption of the pharmaceutical active
agent(s) to the magnetic particles, the coupling can be physical or
chemical in nature or can be based on biological interaction. Such
coupling or adsorption includes e.g. electrostatic, hydrophobic or
hydrophilic interactions, van der Waals interactions, hydrogen bridge
bonds and covalent bonds. Preferred covalent bonds are, for example,
amide, ester, thioester, ether, thioether and disulfide bonds. All
combinations of the interactions mentioned are likewise included. The
coupling between the magnetic particles and pharmaceutical active agent
in the aerosol according to the invention containing magnetic particles
can be established, for example, by a reaction, e.g. a chemical coupling,
of functional groups of the coating of the magnetic particle, as
described above, and functional groups of a vector described here.
Alternatively, the coupling can also be established by using a (homo- or
hetero-bifunctional) linker. Such suitable homo- or hetero-bifunctional
linkers are commercially obtainable. Processes which can be used for a
(chemical) coupling of magnetic particles and pharmaceutical active
agents of the present invention, e.g. using linkers or functional groups
as described above, are well-known in the prior art and are described in
detail, for example, in Bioconjugate Techniques, by Greg T. Hermanson
Academic Press (1 Jan. 1996).

[0042]If the components of the aerosol according to the invention
containing magnetic particles, i.e. magnetic particles and pharmaceutical
active agent(s), are present in coupled form, these can be present both
in combination with a solvent according to the invention and without such
a solvent. In the former case, according to the present invention the
aerosols containing magnetic particles are liquid aerosols, and in the
latter case they are dry aerosols. An aerosol according to the invention
containing magnetic particles in which magnetic particles and
pharmaceutical active agent(s) are present in coupled from in combination
with a solvent as described in the following is particularly preferred.

[0043]The components of the aerosol according to the invention containing
magnetic particles, i.e. the magnetic particle contained therein and the
pharmaceutical active agent, can also be present in the non-coupled form
in the aerosol according to the invention containing magnetic particles.
In order to render possible a directed deposition of the active agent in
such an aerosol according to the invention containing magnetic particles,
a solvent such as is described in the following is preferably employed
together with the magnetic particle and the pharmaceutical active agent.
According to the invention, these are then also liquid aerosols here. In
this case, the transportation is by means of the drop of liquid in which
both (the) magnetic particle(s) and (the) pharmaceutical active agent(s)
are contained, only the magnetic particles contained therein rendering
possible directed transportation and targeted deposition of the active
agent. Such drops of liquid are typically obtained by a nebulizer or an
equivalent device, as described below for the preparation of (liquid)
aerosols according to the invention. An aerosol according to the
invention containing magnetic particles in which magnetic particles and
pharmaceutical active agent(s) are present in non-coupled form, in
combination with a solvent as described in the following, is therefore
likewise particularly preferred.

[0044]A solvent which can be used for an aerosol according to the
invention containing magnetic particles can be an inorganic or organic
solvent. Preferred solvents are ethanol, water and glycerine (glycerol)
or mixtures thereof. Solvents which are suitable according to the present
invention should preferably be tolerated well physiologically by the
organism (mammal, preferably human) to which the aerosol is administered,
i.e. should trigger no side effects, e.g. toxic side effects. Distilled
water is a particularly preferred solvent. Ethanol-water mixtures are
likewise preferred; in this case, the percentage content by weight of
ethanol in these mixtures is preferably in a range of between 5% and 99%
of ethanol, likewise preferably in the range of from 10% to 96% of
ethanol, more preferably between 50% and 92%, most preferably between 69%
and 91% of ethanol.

[0045]The preparation of a solvent-containing aerosol according to the
invention containing magnetic particles with a pharmaceutical active
agent which is not coupled to the magnetic particles can be carried out,
for example, by preparation of drops of liquid comprising a solvent
described above and magnetic particles contained therein and a
pharmaceutical active agent (called formulation according to the
invention in the following). For this, the components mentioned are mixed
and the drops of liquid are generated, for example, with a nebulizer (see
below) or an equivalent device. If the pharmaceutical active agent is
coupled to the magnetic particles, this is carried out before mixing of
the components mentioned, and therefore also before generation of the
drops of liquid from the formulation according to the invention.

[0046]The preparation of aerosols according to the invention containing
magnetic particles without a solvent can be carried out by mixing the
magnetic particle and pharmaceutical active agent components. If
appropriate, for this the pharmaceutical active agent is coupled to the
magnetic particles beforehand and a treatment with a suitable nebulizer,
a propellant gas nebulizer (see below) or an equivalent device is
subsequently carried out. However, the preparation of the aerosols
according to the invention can likewise be carried out by any suitable
process from the prior art.

[0047]The present invention also provides a pharmaceutical composition
which comprises an aerosol according to the invention containing magnetic
particles, and optionally suitable auxiliary substances and/or additives.
The pharmaceutical composition preferably furthermore comprises at least
one (further) solvent, at least one complexing agent and/or at least one
pharmaceutically acceptable acid.

[0048]In connection with the pharmaceutical composition according to the
invention, "auxiliary substances and/or additives" according to the
invention is to be understood as meaning any pharmacologically acceptable
and therapeutically appropriate substance which is not a pharmaceutical
active agent but can be formulated in the pharmaceutical composition
together with the pharmaceutical active agent in order to influence, in
particular to improve, qualitative properties of the pharmaceutical
composition. Preferably, the auxiliary substances and/or additives
display no pharmacological action or, with respect to the intended
therapy, no noticeable or at least no undesirable pharmacological action.
Suitable auxiliary substances and additives are, for example,
pharmacologically harmless salts, for example sodium chloride, flavour
substances, vitamins, e.g. vitamin A or vitamin E, tocopherols or similar
vitamins or provitamins which occur in the human organism, antioxidants,
such as, for example, ascorbic acid, and stabilizers and/or preservatives
for prolonging the use life and storage life of the pharmaceutical
composition, and other conventional auxiliary substances and additives
known in the prior art.

[0049]Preservatives can be employed, for example, in the pharmaceutical
composition according to the invention in order to protect the
pharmaceutical composition from contamination with pathogenic germs.
Suitable preservatives are, in particular, benzalkonium chloride or
benzoic acid or benzoates, such as sodium benzoate, in the concentrations
known from the prior art. The amount of, for example, benzalkonium
chloride added is preferably approximately 1 mg to 50 mg per 100 ml, more
preferably approximately 7 mg to 15 mg per 100 ml, still more preferably
approximately 9 mg to 12 mg per 100 ml of the pharmaceutical composition
according to the invention. However, pharmaceutical compositions
according to the invention which contain no preservatives are
particularly preferred.

[0050]At least one (further) "solvent" of the pharmaceutical composition
according to the invention is to be understood as meaning the solvents
described above in connection with the aerosol according to the invention
containing magnetic particles. If liquid aerosols according to the
invention containing magnetic particles are employed in this context in
the pharmaceutical composition according to the invention, the at least
one (further) "solvent" is at least one solvent in addition to that
contained in the liquid aerosol. In liquid aerosols according to the
invention containing magnetic particles, the at least one (further)
"solvent" is accordingly at least one "solvent". The at least one
(further) "solvent" contained in the pharmaceutical composition according
to the invention can comprise one solvent or any combination of the
solvents described above.

[0051]In connection with the pharmaceutical composition according to the
invention, "complexing agents" are to be understood as meaning molecules
which are suitable for entering into complex bonds. Preferably, cations,
particularly preferably metallic cations, are complexed according to the
invention by these compounds. Preferred complexing agents are edetic acid
(EDTA, ethylenediaminetetraacetate) or a known salt thereof, e.g. sodium
EDTA or disodium EDTA. Preferably, sodium edetate
(ethylenediaminetetraacetic acid disodium salt), optionally in the form
of its hydrates, particularly preferably in the form of its dihydrate, is
employed. The concentration of the complexing agents employed is
preferably in a range of from approximately 1 mg to 100 mg per 100 ml,
more preferably in a range of from approximately 5 mg to 50 mg per 100
ml, more preferably from approximately 6 mg to 30 mg per 100 ml, most
preferably from approximately 7 mg to 20 mg per 100 ml of the
pharmaceutical composition according to the invention.

[0052]Pharmaceutically acceptable inorganic or organic acids are used, in
particular, to adjust the pH of the pharmaceutical composition according
to the invention, this pH preferably being in a range of from
approximately 3.0 to approximately 8.5, preferably between 3.5 and 6.0,
particularly preferably between 4.0 and 7.0. The pH is very particularly
preferably about 7.0. Pharmaceutically acceptable inorganic or organic
acids can in principle be selected from all suitable acids or bases. In
connection with the pharmaceutical composition according to the
invention, examples of preferred (pharmaceutically acceptable) inorganic
acids can be selected from the group consisting of hydrochloric acid,
hydrobromic acid, nitric acid, sulfuric acid and phosphoric acid,
hydrochloric acid and sulfuric acid being preferred in particular. In
connection with the pharmaceutical composition according to the
invention, examples of particularly suitable (pharmaceutically
acceptable) organic acids can be selected from the group consisting of
malic acid, tartaric acid, maleic acid, succinic acid, acetic acid,
formic acid and propionic acid, and particularly preferably ascorbic
acid, fumaric acid and citric acid. Mixtures of the acids mentioned can
optionally also be employed, in particular of acids which, in addition to
their acidification properties, also have other properties, e.g. in use
as flavour substances or antioxidants, such as, for example, citric acid
or ascorbic acid.

[0053]For exact adjustment of the pH described above for the
pharmaceutical composition according to the invention, pharmaceutically
acceptable bases can furthermore optionally also be employed. Suitable
bases in this connection are alkali metal hydroxides, alkali metal
carbonates and alkali metal ions, preferably sodium. It is to be ensured
that the salts resulting from such bases, which are then contained in the
pharmaceutical composition according to the invention, are
pharmaceutically acceptable with the above-mentioned acid(s). The
combination of the particular suitable acids and bases for adjustment of
the pH of the pharmaceutical composition according to the invention can
be varied as required and according to the requirements of the
composition to be prepared. A corresponding choice or combination of such
acids and bases as described above can easily be made by a person skilled
in the art.

[0054]The preparation of a pharmaceutical composition according to the
invention can be carried out analogously to the preparation described
above for the aerosols according to the invention. Further components
optionally contained in the pharmaceutical composition according to the
invention, i.e. auxiliary substances and/or additives, the solvent
preferably contained therein, complexing agents and/or at least one
pharmaceutically acceptable acid, can be added in the aerosol preparation
process described above by processes known in the prior art.
Alternatively, the pharmaceutical composition according to the invention
can be achieved by any process known in the prior art.

[0055]The dosage of the components described above in the pharmaceutical
composition according to the invention is subject to various factors, for
example the nature of the treatment, the disease, the condition of the
organism (mammal, preferably human), the age of the sick organism to
which the pharmaceutical composition according to the invention is
administered, the nature of the active agent etc. Such parameters are
known to the person skilled in the art and the determination of the
dosages is subject to his general technical knowledge.

[0056]The aerosol according to the invention containing magnetic particles
and the pharmaceutical composition according to the invention serve in
particular for nasal or oral administration. A preferred embodiment
therefore relates to an aerosol according to the invention containing
magnetic particles or a pharmaceutical composition according to the
invention for nasal or oral administration. The administration is
preferably carried out via a nebulizer. A further preferred embodiment
therefore relates to an aerosol according to the invention containing
magnetic particles or a pharmaceutical composition according to the
invention for administration via a nebulizer.

[0057]A "nebulizer" in the context of the present invention is any
standard nebulizer suitable for medicinal aerosols. The term "nebulizer"
is to be understood as being synonymous with the term "inhaler".
Nebulizers are conventionally used to prepare and to administer liquid
and/or dry aerosols according to the invention containing magnetic
particles, e.g. based on solvents (as already described above). For this,
the formulations according to the invention are conventionally fed to a
nebulizer in order to prepare from this the preferably propellant
gas-free aerosols according to the invention containing magnetic
particles. For this, the nebulizer typically sprays a defined volume of
the formulation using high pressures through small jets, in order to
generate an inhalable aerosol according to the invention containing
magnetic particles in this way. Nebulizers which are particularly
suitable here are those which can nebulize a small amount of a liquid
formulation according to the invention in the therapeutically necessary
dosage within a few seconds into an aerosol suitable for therapeutic
inhalation. Such nebulizers are suitable in particular for propellant
gas-free administration of the (liquid) aerosols according to the
invention containing magnetic particles or pharmaceutical compositions
according to the invention. Such a nebulizer is described e.g. in the
international patent applications WO 91/14468 and WO 97/12687. In such a
nebulizer, a medicament solution is converted by means of a high pressure
of up to 600 bar into a medicinal aerosol which is suitable for
administration to the respiratory tract and lung and is sprayed. A
specific jet such as is described, for example, in WO 94/07607 or WO
99/16530, is used for nebulizing such a solution.

[0058]Suitable nebulizers for nasal or oral administration of the dry
aerosols according to the invention containing magnetic particles, i.e.
aerosols according to the invention containing magnetic particles without
a solvent, are typically inhalation apparatuses or nebulizers driven by
propellant gas. Propellant gases here can be, for example, CFCs or HFCs.
In this respect, reference is made to the publication "Theorie und Praxis
der Inhalationstherapie [Theory and Practice of Inhalation Therapy],
pages 31 to 70, Arcis Verlag (2000), in which a detailed description of
nebulizers which can be used and methods of use thereof is/are disclosed.

[0059]Further examples of suitable nebulizers for nasal or oral
administration of the aerosols according to the invention containing
magnetic particles are jet nebulizers driven by compressed air (e.g. PARI
LC plus, PARI GmbH, Starnberg, Germany), Venturi jet nebulizers, jet
nebulizers driven by water vapour or ultrasound nebulizers (e.g.
AeronebLab, Aerogen, Inc., Stierlin Court, Canada; eFLOW, PARI GmbH,
Starnberg, Germany). Nebulizers of a size which can be carried by the
patient (human), e.g. the Respimat@, as described in WO 97/12687, are
likewise suitable. A detailed description of suitable nebulizers is also
to be found in "Theorie und Praxis der Inhalationstherapie [Theory and
Practice of Inhalation Therapy]", pages 31 to 70, Arcis Verlag (2000).

[0060]The present invention therefore also additionally provides a device
for nebulizing and/or administration of the aerosol according to the
invention containing magnetic particles, comprising a nebulizer as
described above filled with an aerosol according to the invention
containing magnetic particles. The device according to the invention can
be configured as a device having a replaceable reservoir of aerosol
according to the invention containing magnetic particles. Alternatively,
the device according to the invention can be a disposable device. The
device according to the invention can furthermore be mobile or
stationary. If the device according to the invention is mobile, the
weight is preferably not more than 2 kg, particularly preferably not more
than 1 kg, still more preferably not more than 500 g. If the device
according to the invention is stationary in configuration, the device
preferably comprises a replaceable reservoir of aerosol according to the
invention containing magnetic particles.

[0061]The present invention also provides the use of an aerosol according
to the invention containing magnetic particles or of a pharmaceutical
composition according to the invention for prophylaxis and/or therapy of
diseases of the respiratory tract and/or lungs, in particular
inflammatory and/or obstructive diseases of the respiratory tract and/or
lungs, for example melanomas or malignant melanomas of the respiratory
tract, in particular the lung, lung cancer, lung tumours, lung
carcinomas, small cell lung carcinomas, throat cancer, bronchial
carcinomas, larynx cancer, head/neck tumours, tongue cancer, sarcomas and
blastomas in the region or vicinity of the respiratory tract, in
particular the lung, as well as asthma and COPD (chronic obstructive
pulmonary disease), lung emphysema, chronic bronchitis, pneumonia and
hereditary diseases, for example mucoviscidosis, human surfactant protein
B deficiency and α1-antitrypsin deficiency, and for therapy
after a lung transplant and for pulmonary vaccination and for
anti-infective therapy of the lung (in cases of bacterial (e.g.
Bronchiolitis obliterans) or viral colonization of the lung).

[0062]The present invention also provides the use of an aerosol according
to the invention containing magnetic particles or optionally of a
pharmaceutical composition according to the invention for the preparation
of a medicament or a device for prophylaxis and/or therapy of diseases of
the respiratory tract and/or lungs, in particular inflammatory and/or
obstructive diseases of the respiratory tract and/or lungs, for example
melanomas or malignant melanomas of the respiratory tract, in particular
the lung, lung cancer, lung tumours, lung carcinomas, small cell lung
carcinomas, throat cancer, bronchial carcinomas, larynx cancer, head/neck
tumours, tongue cancer, sarcomas and blastomas in the region or vicinity
of the respiratory tract, in particular the lung, as well as asthma,
COPD, lung emphysema, chronic bronchitis, pneumonia and hereditary
diseases, for example mucoviscidosis, human surfactant protein B
deficiency and α1-antitrypsin deficiency, and for therapy
after a lung transplant and for pulmonary vaccination and for
anti-infective therapy of the lung (in cases of bacterial (e.g.
Bronchiolitis obliterans) or viral colonization of the lung).

[0063]The present invention also provides the use of an aerosol according
to the invention containing magnetic particles or of a pharmaceutical
composition according to the invention in diagnostics. The present
invention alternatively provides the use of an aerosol according to the
invention containing magnetic particles or of a pharmaceutical
composition according to the invention for the production of a device in
diagnostics. For example, the diagnostic agent can be a contrast medium,
including e.g. iodine-containing (water-soluble) x-ray-positive contrast
media; iodine-containing (water-insoluble) x-ray-positive contrast media;
barium containing (water-insoluble) x-ray-positive contrast media;
x-ray-negative contrast media; gadolinium compounds, such as e.g.
gadolinium DTPA; technetium compounds, such as e.g. compounds containing
technetium-99m, for example phosphonates labelled with technetium-99m,
Tc-99m-MDP=Tc-99m-methylene-diphosphonate, Tc-99m-tetrofosmin;
superparamagnetic iron oxide particles as described above; contrast media
which can pass through membranes; enzyme-specific NMR probes, e.g. with
paramagnetic dinuclear complexes with lanthanoid ions based on cyclic
polyaminopolyacetic acids; glucose contrast media, for example for use in
PET (positron emission tomography), including e.g. glucose contrast media
coupled to a labelled substance which renders possible external location,
e.g. using isotopes which emit positrons, e.g. 18-fluorine, such as,
for example, fluoro-2-deoxy-2-D-glucose labelled with 18-fluorine
(18FDG), etc. The diagnostic agent can furthermore comprise peptides,
proteins, nucleic acids, antibodies, in particular detectable antibodies,
low molecular weight detectable compounds, etc. For diagnostics, the
aerosol according to the invention containing magnetic particles or a
pharmaceutical composition according to the invention can be introduced
by the directed transportation described into defined regions of the
respiratory tract, in particular the lung, and can function as a
diagnostic agent in these regions. By this means, structural changes to
the lung which indicate a pathological change can be detected and imaged
early on, and therefore facilitate the diagnosis of a potential disease.
Such a diagnostic agent can be used in particular for imaging lung
tumours or lung emphysemas (imaging MRI, CT, determination of the extent
of a lung emphysema).

[0064]A preferred embodiment of the present invention relates to the use
of an aerosol according to the invention containing magnetic particles or
of a pharmaceutical composition according to the invention for directed
deposition of aerosol in the respiratory tract or in the lung of an
organism (mammal, preferably human) and the use of an aerosol according
to the invention containing magnetic particles or of a pharmaceutical
composition according to the invention preferably for nasal or oral
administration, preferably via a nebulizer.

[0065]The present invention also provides a method for inhalatory use of
an aerosol according to the invention containing magnetic particles or of
a pharmaceutical composition according to the invention, wherein the
method comprises the preparation of an aerosol according to the invention
containing magnetic particles or of a pharmaceutical composition
according to the invention and the administration of this to an organism
(mammal, preferably human). The preparation and administration are
preferably carried out as described above.

[0066]Preferably, the method for inhalatory use of an aerosol according to
the invention containing magnetic particles is carried out using a
magnetic field which is applied externally as described above and is
permanently, or still more preferably is not permanently active.
Particularly preferably, during the administration the externally applied
magnetic field is active only during the period of the resting phases
between inhalation and exhalation or between exhalation and inhalation.
Still more preferably, the guiding of the magnetic field can take place
dynamically in coordination with the breathing of the patient as a
function of the breathing rhythm of the patient, so that during the in-
and exhalation of the patient, no magnetic field is applied in the region
to be treated, but in the resting phases a magnetic field is applied
there and only then does a deposition of the aerosol according to the
invention containing magnetic particles on the surface of the respiratory
tract take place. Preferably, for this the air above the chosen surface
of the respiratory tract is saturated by administration of the aerosol
according to the invention containing magnetic particles such that within
several activations of the magnetic field(s) by the magnet(s) a
significant deposition of the transported active agent or of the aerosol
according to the invention containing magnetic particles on the chosen
surface of the respiratory tract is rendered possible. Control of the
magnet can be rendered possible, for example, by an electric circuit
which triggers a signal in the nebulizer or inhaler at the start and end
of the inhalation or exhalation by the patient, by which in turn the
magnetic field of a magnet as described here is switched on or off.

[0067]The present invention also provides a kit comprising an aerosol
according to the invention containing magnetic particles or a
pharmaceutical composition according to the invention, an external magnet
which generates a magnetic field and a nebulizer.

[0068]All the references given in the description of the present invention
are included in their full scope in the present invention. The invention
is explained in more detail in the following with the aid of figures and
examples. It is not intended to limit the present invention to these.

[0069]FIG. 1 shows the various mechanisms of deposition of aerosol
particles in the lung. The hypothetical deposition on the inner wall of
the air cavity (respiratory tract epithelium) of various particles is
shown as a function of their diameter and the consequent deposition
mechanism

[0070]FIG. 2 shows the normal local deposition of aerosol particles in the
lung during calm nasal breathing as a function of their diameter. Studies
with respect to local deposition have shown that the particle diameter
has a decisive influence on local deposition. [0071]It can be seen from
FIG. 2 that inhaled particles having a diameter of between 0.1 to 1 μm
overall are deposited only minimally in the lung. On the other hand, the
deposition of particles of larger and smaller diameter increases overall.
It can likewise be seen from FIG. 2 that particles having a diameter of
0.01 μm are deposited to the extent of approx. 80% and of 5 μm are
deposited to the extent of approx. 100%. The increasing deposition of
larger particles is to be attributed above all to the increasing
deposition by sedimentation and impaction as the particle size increases.
In contrast, the deposition of small particles of <0.1 μm increases
due to deposition by diffusion. The total deposition during calm nasal
breathing is comparable in rats and in humans. [0072]It has been found,
according to the invention, that it was possible to increase considerably
the deposition of particles by the magnetic field-guided deposition
according to the invention (see also FIG. 15). [0073]With respect to the
local deposition of the particles, FIG. 2 shows that this also depends
greatly on the particle diameter. Extrathoracic deposition takes place
chiefly in the nose and is only slightly lower than the total deposition.
In particular, particles having a diameter larger than 1 μm or smaller
than 0.05 μm are already filtered in the nose. The remaining
proportion of the inhaled aerosol is mostly deposited in the alveolar
region. The aerosol proportion deposited in the tracheobronchial region
(trachea and respiratory tract) is relatively low, and reaches up to 15%
for very small particles, while particles of >3 μm are deposited to
the extent of only 1 to 3%. In the alveolar region, approx. 15% of the
inhaled dose of particles of diameter of <0.1 μm is deposited. For
particles having a diameter of from 0.1 to 3 μm, the proportion
deposited varies between 5 to 10%. Deposition is almost negligible for
particles of >5 μM. Overall, it can be said that tracheobronchial
deposition is very similar between rats and humans (see Schulz, H. &
Muhle, H. 323-345 (Academic Press, 2000)).

[0074]According to the invention, it has been found (see also FIG. 9) that
by the magnetic field-guided deposition according to the invention, the
deposition of magnetic particles can be effected and also increased in a
targeted manner in regions in which normally only a low deposition takes
place, as described above, for example, for the tracheobronchial and the
alveolar region

[0075]FIG. 3 shows the deposition of particles using a modern jet
nebulizer operated with compressed air (PARI IS-2). Of a total inhaled
aerosol dose (exhalation, extrabronchial and intrabronchial), 52% is
exhaled again (see Kohler, D. & Fischer, F. Theorie und Praxis der
Inhalationstherapie [Theory and Practice of Inhalation Therapy] (Arcis
Verlag GmbH, Munich, 2000). These results illustrate that a large
proportion of the inhaled aerosol dose is exhaled again.

[0076]This exhaled proportion is minimized considerably with the present
invention by the magnetic field-guided deposition of aerosol. It can be
seen from FIG. 3 that in total only 33% of the inhaled dose is deposited
in the lung.

[0077]The results from experiments of the invention (shown in FIGS. 5 and
6) show that an approx. 2.2- to 2.5-fold increase in this inhaled dose is
deposited when the influence according to the invention of an external
magnetic field is present. Since in the abovementioned experiments the
breathing conditions with and without a magnetic field were identical, on
application of the results to the data from FIG. 3 it emerges that the
aerosol dose deposited in the lung can be increased from 33% to approx.
82.5% with the aerosols according to the invention.

[0078]FIG. 4 shows the deposition pattern of conventionally administered
aerosols using inhalation methods which correspond to the prior art. As
can be seen, the inhaled aerosol is distributed homogeneously throughout
the entire lung. A directed guiding of the aerosol into defined regions
of the lung is not possible with the methods known hitherto.

[0079]FIG. 5 shows a diagram of a cross-section through the mucus layer
and the respiratory epithelium. This figure gives an overview of
conditions of in vivo transfer of pharmaceutical active agents (and
magnetic particles) of the present invention. After topical application
of, for example, gene vectors via the respiratory tract, extracellular
and intracellular barriers are to be overcome before the DNA enters into
the cell nucleus. The lung surfactant film, mucus layer, proteins
contained therein, such as e.g. nucleases (DNase), form a first barrier
in this context. Further physical barriers are the cilia and glycocalix
of the cell surface. After intracellular uptake of gene vectors, the DNA
must be protected from lysosomal degradation and transported to the cell
nucleus. The cell nucleus pores of the nucleus membrane are a further
obstacle to successful transportation. The mucus layer per se is a
barrier which is difficult to penetrate. Diffusion of particles of
>500 nm through the mucus layer therefore virtually stops (see
Sanders, N. N., De Smedt, S. C. & Demeester, J. The physical properties
of biogels and their permeability for macromolecular drugs and colloidal
drug carriers [Review]. Journal of Pharmaceutical Sciences 89, 835-849
(2000)).

[0080]FIG. 6 shows the cells of the lung lying underneath the mucus layer
(as shown in the cross-section from FIG. 5). FIG. 6A shows the cells
after administration of an aerosol according to the invention containing
magnetic particles under the influence of a magnetic field, FIG. 6B shows
the cells after administration of the corresponding aerosol according to
the invention containing magnetic particles without the influence of a
magnetic field. As can be seen in FIG. 6A, the magnetic particles adhere
to the cell surface and they are to be seen as black deposits. On the
other hand, no deposits are detectable on the cell surface in FIG. 6B.
With this study result it was possible to demonstrate that the aerosols
according to the invention containing magnetic particles or the magnetic
particles and pharmaceutical active agents (which are either coupled to
the magnetic particles or contained with them in a drop of liquid, as
described above) contained therein are transported in a directed manner
through the mucus layer, whereas this is not the case without the
application according to the invention of an external magnetic field.

[0081]FIG. 7 shows a diagram of the concept according to the invention of
the magnetic field-guided administration of aerosol. An aerosol according
to the invention containing magnetic particles is generated with a
commercially obtainable nebulizer (PARI, Starnberg, Germany). The drops
of liquid containing magnetic particles formed here are inhaled by the
patient and guided by the externally applied magnetic field directly into
the desired regions of the lung of the patient.

[0082]FIG. 8 shows a diagram of an animal study on mice of the concept
according to the invention of the magnetic field-guided administration of
aerosol. Aerosols containing magnetic particles were guided in a targeted
manner into defined regions of the lung of a mouse under the influence of
an applied magnetic field.

[0083]FIG. 9 shows the study set-up of an animal study on mice of the
concept according to the invention of the magnetic field-guided
administration of aerosol. In the mouse model, the thorax of the mice was
opened, so that on the one hand it was possible to expose and intubate
the trachea, and on the other hand it was possible to apply an external
magnetic field to a defined region of the lung. The mouse was respirated
(the flexiVent system from SCIREQ is shown) during administration of the
aerosol, and the aerosol was introduced through a connected nebulizer
into the respiration system (symbolized by the red arrow). The nebulizer
was synchronized with the respiration frequency, so that aerosol was
administered into the mouse lung only during inspiration. [0084]The
general study set-up of the intubated mouse connected to the respirator
and the nebulizer is shown in FIG. 9A. In FIG. 9B, the intubated mouse is
shown, fixed under the electromagnet, which was used as the external
magnetic field. The intubated mouse with the opened thorax (without the
magnet) can be seen in FIG. 9C. FIG. 9D shows the iron tip (pole shoe) of
the electromagnet placed on the right mouse lung. The mouse was fixed
under the electromagnet during administration of the aerosol such that
the iron tip of the electromagnet did not come into contact with the left
mouse lung. The precise course of the study is disclosed in Example 1.

[0085]FIG. 10 shows the histological evaluation of an animal study on mice
of the magnetic field-guided aerosol administration according to the
invention. The distribution of superparamagnetic iron oxide nanoparticles
after administration of aerosol into the lung tissue of a mouse is shown.
A magnetic field was applied to the right lung during administration of
the aerosol. The superparamagnetic iron oxide nanoparticles appear as a
brown colouration and can be seen in FIG. 10 as dark regions in the
image. [0086]Longitudinal sections through the mouse lung are shown in
the first row of FIG. 10. The pointwise accumulation of the
superparamagnetic iron oxide nanoparticles at the position of the pole
shoe of the electromagnet can be clearly seen (left-hand image). In
contrast, no pointwise accumulation of the superparamagnetic iron oxide
nanoparticles is to be seen in the left lung, but a homogeneous and
diffuse distribution of the superparamagnetic iron oxide particles over
the lung (middle image). A control lung in which as expected no brown
colouration of the lung is to be observed is also shown (right-hand
image). In all the control lungs of FIG. 10, no superparamagnetic iron
oxide nanoparticles were administered as an aerosol. [0087]The second row
of FIG. 10 shows marked sections from the lung sections of the first row
in magnification. The accumulation of the superparamagnetic iron oxide
nanoparticles induced in the right mouse lung by the magnetic field can
be clearly seen (left-hand image), while this is not observed in the left
mouse lung (middle image). Control lungs in which as expected no
accumulation of superparamagnetic iron oxide nanoparticles is to be
observed are likewise shown (right-hand images). [0088]In the third and
fourth row of FIG. 10, marked sections of the enlargements of the second
row are shown in turn. A clear accumulation of the superparamagnetic iron
oxide nanoparticles in the lung is to be seen only in the regions in
which the pole shoe of the electromagnet was placed (left-hand images).
Control lungs in which as expected no accumulation of superparamagnetic
iron oxide nanoparticles is to be observed are likewise shown (right-hand
images). [0089]These results shown in FIG. 10 illustrate that it is
possible to guide an aerosol according to the invention containing
magnetic particles successfully in a controlled manner into defined
regions of the lung by means of externally applied magnetic fields. The
invention therefore represents a substantial step in the direction of a
completely novel technology for administration of aerosol and for
targeted therapy of numerous diseases of the respiratory tract and/or
lungs.

[0090]FIG. 11 shows the quantitative evaluation of an animal study on mice
of the magnetic field-guided aerosol administration according to the
invention. In this study, the thorax of the mouse was opened and an
electromagnet was placed on the right lung. In FIG. 11, the results of
the left lung (without a magnetic field applied) are compared with those
of the right lung (with a magnetic field applied). [0091]In FIG. 11,
the amount of the quantity of superparamagnetic iron oxide nanoparticles
deposited in the lungs of the mouse under the influence of a magnetic
field applied only to the right mouse lung is shown. Aerosol
administration of superparamagnetic iron oxide nanoparticles in the
absence of a magnetic field leads to a uniform distribution between the
right and left mouse lung (white columns). In contrast, application of a
magnetic field to the right mouse lung causes an approx. 8-fold
accumulation compared with the left mouse lung (black columns).
[0092]Viewed overall, the total deposition of superparamagnetic iron
oxide nanoparticles in the mouse lung is at least 2.8 times higher under
the influence of the magnetic field than in the absence of the magnetic
field. According to the invention, the directed deposition of aerosols of
the present invention containing magnetic particles has also been
demonstrated by these studies and results.

[0093]FIG. 12 shows the quantitative evaluation of an animal study on mice
of the magnetic field-guided aerosol administration according to the
invention. In this study, the thorax of the mouse was closed and an
electromagnet was placed on the right lung. FIG. 12 compares the results
of the left lung (without a magnetic field applied) to those of the right
lung (with a magnetic field applied). [0094]The background of this
study was also use in humans, where the magnetic field-guided
administration of aerosol has to take place with the thorax closed. As
can be seen in FIG. 12, with the thorax closed the influence of the
magnetic field applied to the right lung also results in an at least
2.5-fold increase in the deposition of the superparamagnetic iron oxide
nanoparticles, and therefore virtually corresponds to the results of the
administration of aerosol with the thorax opened (shown in FIG. 11). It
has thus been demonstrated that the present invention can be used in
human aerosol therapy and under the associated conditions (closed
thorax).

[0095]FIG. 13 shows a further quantitative evaluation of an animal study
on mice of the magnetic field-guided aerosol administration according to
the invention. In this study, plasmid DNA (pCMVLuc) (as the active agent)
was formulated with superparamagnetic iron oxide nanoparticles and
administered as an aerosol according to the invention. In this study
also, the thorax of the mouse was closed and an electromagnet was placed
on the right lung. FIG. 13 compares the results of the left lung (without
a magnetic field applied) to those of the right lung (with a magnetic
field applied). [0096]The background of this study was also here use in
humans, where the magnetic field-guided administration of aerosol has to
take place with the thorax closed. As can be seen from FIG. 13, with the
thorax closed the influence of the magnetic field applied to the right
lung also results in an at least 2.2-fold increase in the deposition of
the plasmid DNA administered, which was formulated with superparamagnetic
iron oxide nanoparticles, and therefore virtually corresponds to the
results of the administration of aerosol with the thorax opened (shown in
FIG. 10). It has thus been demonstrated again that the present invention
can be used in human aerosol therapy and under the associated conditions
(closed thorax).

[0097]FIG. 14 shows a diagram of a study set-up for investigating the
magnetic field-induced deflection of the aerosol. [0098]A test model
was established for magnetic field-guided pharmaceutical active agent
transfer according to the invention in aerosol therapy. A diagram of the
study set-up of the test model is shown in FIG. 14. The test model is
characterized by the following features: A standard nebulizer (PARI LC
plus; made available by the partner company PARI GmbH, Starnberg) was
connected to a 60 cm long hose of plastic having an internal diameter of
0.7 cm, and various magnets were positioned on the outer wall of the hose
at a distance of 30 cm from the nebulizer. The nebulizing conditions
chosen correspond to the diameter and the air speed such as are to be
found in the 1st-2nd bronchial branching of the human lung. Aqueous
formulations according to the invention containing superparamagnetic iron
oxide nanoparticles were nebulized into the hose for defined times and
the magnetic field-dependent deflection efficiency was quantified by
measuring the hose area containing iron oxide nanoparticles.

[0099]FIG. 15 shows the magnetic field-induced aerosol deflection of
aerosols containing superparamagnetic iron oxide particles. [0100]To
illustrate the magnetic field-induced deflection of aerosols according to
the invention containing iron oxide particles, the deposition of the iron
oxide particles on the inner wall of the hose of the test model shown in
FIG. 14 is shown by way of example in FIG. 15. In this study, a bar
magnet mounted with an iron tip was used. The nebulizing parameters are
stated. The deposition of aerosol in the hose without application of the
magnetic field is shown as a control in the upper photo of FIG. 15. It
can be clearly seen that the iron oxide particles are deposited uniformly
on the base of the hose if no magnetic field is applied. In contrast, the
accumulation of the iron oxide particles at the iron tip of the permanent
magnet on application of the magnetic field can be clearly seen. It can
likewise be clearly seen that the flight path of the aerosol particles
changes as a function of the magnetic field, since no deposition of the
iron particles is to be seen on the base of the hose underneath the iron
tip of the magnet.

[0101]FIG. 16 shows the magnetic field-induced aerosol deflection of
aerosols according to the invention containing superparamagnetic iron
oxide particles. [0102]Various magnet arrangements were investigated by
means of the test model shown in FIG. 14. It emerged above all that
generation of magnetic field gradients by targeted placing of pieces of
iron (e.g. paper clips) in the magnetic field greatly increased the
deposition of the aerosols. The most efficient arrangement was achieved
by placing a permanent magnet in each case above and below the hose. The
result is shown in FIG. 16.

[0103]FIG. 17 shows the further parameter of the field strength, which has
an influence on the deflection properties of aerosols according to the
invention containing superparamagnetic iron oxide particles. The
investigations were carried out with the aid of the test model shown in
FIG. 14. [0104]It can be clearly seen that the deposition of the
aerosols containing superparamagnetic iron oxide particles decreases
greatly with the magnetic field strength. As expected, the greater the
magnetic field, the greater the deflection of the aerosol.

[0105]FIG. 18 shows the further parameter of the magnetic particle
diameter, which has an influence on the deflection properties of aerosols
according to the invention containing superparamagnetic iron oxide
particles. The investigations were carried out with the aid of the test
model shown in FIG. 14. [0106]It can be seen that the deposition of the
aerosol depends on the size of the diameter of the superparamagnetic iron
oxide particles. Interestingly, the deflection of small particles of 50
nm in diameter is entirely more effective that that of larger particles
of more than 800 nm at the same iron concentration. The magnetic
particles used according to the invention having a diameter of up to 800
nanometers therefore show significantly better results for transportation
of aerosol than magnetic particles having a diameter larger than 800
nanometers. The aerosols according to the invention containing magnetic
particles are therefore excellently suitable for use in aerosol therapy.

[0107]FIG. 19 shows the further parameter of the iron concentration, which
has an influence on the deflection properties of aerosols according to
the invention containing superparamagnetic iron oxide particles. The
investigations were carried out with the aid of the test model shown in
FIG. 14. [0108]A further parameter which has a great influence on the
deflection properties is the concentration of the nebulized
superparamagnetic iron oxide particles in the aerosol solution. In FIG.
19, this is shown by way of example for particles of 100 nm in size. It
can be seen that a minimum concentration of 12.5 mg/ml of solution must
be used, so that the aerosols containing superparamagnetic iron oxide
particles can be deflected.

[0110]FIG. 21 describes the results of in vivo studies with BALB/c mice
(see Example 4). For this, two permanent magnets were fixed on the thorax
of BALB/c mice and an aerosol according to the invention containing
magnetic particles was administered. 24 hours after administration of the
aerosol according to the invention containing magnetic particles, the
mice (n=3) were sacrificed and the amount of plasmid DNA in the lungs was
determined by means of a real-time PCR. As can be seen in FIG. 21, the
presence of the permanent magnets on the thorax led to a 2.5-fold higher
deposition of plasmid DNA in the lung.

[0111]FIG. 22 describes the results of in vivo studies with anaesthetized
domestic pigs (see Example 5). For this, an aerosol according to the
invention containing magnetic particles was administered to two BALB/c
anaesthetized domestic pigs. On the first pig, a permanent magnet was
placed on the right side of the breast, and no magnet was placed on the
second pig during the administration. After the administration of the
aerosol, the pigs were sacrificed and the lungs removed. The magnetic
particles in the ventral lung regions were quantified by means of
magnetorelaxometry. As can be seen in FIG. 22, the number of magnetic
particles in the right lung by means of a magnetic field is 5 times
higher than without a magnetic field.

EXAMPLES

Example 1

Administration of an Aerosol According to the Invention Containing
Magnetic Particles

[0112]BALB/c mice were anaesthetized with an intraperitoneal injection of
pentobarbital. After the corneal reflex had stopped, the animals were
intubated or connected to the Flexivent system with a nebulizer via a
tracheotomy. The mice were respirated under volume control with a
breathing frequency of f=120 min-1 and a tidal volume of TV=10
μl/g at a PEEP of 4 cm H2O. Before the start of the inhalation,
the lung impedance was measured as the starting value
(pertubation+snapshot).

[0113]The pole shoe (iron tip) of an electromagnet was placed over the
right mouse lung and operated with a current intensity of I=10 A (1 T).
0.7 ml of an aqueous formulation according to the invention with
superparamagnetic iron oxide nanoparticles (fluidMAG-PEI 50 nm, c=6.25
mg/ml) was administered in 10 s intervals (n=20, output rate=200
μl/min). At the end of the experiment, the mouse lungs were removed
and the right and left mouse lung were deep-frozen in liquid nitrogen
separately from one another. The content of magnetic particles in the
lungs was quantified by means of magnetorelaxometry (Lange J, et al.,
Magnetorelaxometry, a new binding specific detection method based on
magnetic nanoparticles, JOURNAL OF MAGNETISM AND MAGNETIC MATERIALS 252
(1-3): 381-383 November 2002). The study group size was 4 mice (3 mice
for quantification of the magnetic particle content of the lungs by means
of magnetorelaxometry and one mouse for the histological processing
(cryosections)).

[0114]Superparamagnetic iron oxide nanoparticles which are coated with the
cationic polymer polyethylenimine (50 nm) (fluidMAG-PEI, Chemicell,
Berlin, Germany) are diluted in distilled water to a final iron
concentration of 12.5 mg/ml. The volume is 4 ml. Plasmid DNA (pCMVLuc) is
diluted with distilled water to a concentration of 0.625 mg/ml (final
volume 4 ml). This results in a total dose of 5 mg of DNA. The plasmid
DNA solution is pipetted into the iron oxide nanoparticle solution and
the components are mixed thoroughly by pipetting up and down. The
resulting pH of the solution is adjusted to pH=7.0. The solution is
incubated at room temperature for 10 min before administration by
nebulizing. It is to be ensured that the electrolyte concentration is
kept as low as possible, in order to prevent a salt-induced aggregation
of the iron oxide nanoparticles. For this reason, no electrolytes but
nonionic substances, such as, for example, glucose or glycerol, should be
used for isotonic adjustment of solutions. The duration of the nebulizing
depends on the type of nebulizer and for a PARI BOY (PARI GmbH,
Starnberg, Germany) is approx. 15 min for a 4 ml volume with an ejection
rate of 5-6 l/min. This results in a nebulizing time of approx. 30 min.
The magnetic field applied is adjusted to a magnetic field gradient of at
least 10 T/m and is placed at the desired position of the lung during the
nebulizing.

[0115]Doxycycline HCl is dissolved in a 20% ethanolic aqueous solution
with a concentration of 16 mg/ml in a volume of 4 ml. 4 ml of a solution
of superparamagnetic iron oxide nanoparticles (c=12.5 mg/ml, 50 nm,
fluidMAG-PEI, Chemicell, Berlin, Germany) are pipetted into this
solution. The solution is nebulized under the conditions mentioned in
Example 2.

[0116]Paclitaxel is dissolved in a mixture of ethanol and polyethylene
glycol 200 (PEG-200) with a concentration of 75 mg/ml in a volume of 4
ml. 4 ml of a solution of superparamagnetic iron oxide nanoparticles
(c=12.5 mg/ml, 50 nm, fluidMAG-PEI, Chemicell, Berlin, Germany) are
pipetted into this solution. The solution is nebulized under the
conditions mentioned in Example 2.

[0117]Dilauroylphosphatidylcholine and paclitaxel are dissolved in a ratio
of 10:1 (w/w) in t-butanol and the solution is frozen at -70° C.
and lyophilized. The lyophilisate is stored at -20° C. until used.
Before use, the lyophilisate is reconstituted with sterile distilled
water and vortexed until multilamellar liposomes are formed. The final
concentration of paclitaxel is 10 mg/ml in a total volume of 4 ml. 4 ml
of a solution of superparamagnetic iron oxide nanoparticles (c=12.5
mg/ml, 50 nm, fluidMAG-PEI, Chemicell, Berlin, Germany) are pipetted into
this solution. The solution is nebulized under the conditions mentioned
in Example 2.

Example 4

In Vivo Studies with BALB/c Mice

[0118]Two permanent magnets (NeoFeBr, 10×10×10 mm; 500 mT)
were fixed on the thorax of BALB/c mice (Elevage Janvier, Route Des
Chenes Secs, Le Genest Saint Isle, 53940 France) with tissue adhesive.
The mice were placed in a Plexiglas box which was divided into six
chambers of equal size of wire permeable to aerosol. A mouse with or
without magnet was placed alternately in the individual chambers. For
preparation of the aerosol according to the invention containing magnetic
particles, plasmid DNA which codes for the luciferase gene was diluted
with distilled water to a volume of 5 ml (c=0.2 mg/ml). Superparamagnetic
iron oxide nanoparticles (fluidMAG-PEI, 50 nm) were then diluted with
distilled water to a volume of 5 ml (c=3 mg/ml), the DNA solution was
pipetted rapidly into the nanoparticle solution and the components were
mixed thoroughly by pipetting up and down several times. Thereafter, the
solution was administered to the mice in the Plexiglas box by means of a
nebulizer. The solution is nebulized under the conditions mentioned in
Example 2. The general procedure for administration of aerosol is
described in the publication of Rudolph et al. (2005) (Rudolph C, Ortiz
A, Schillinger U, Jauernig J, Plank C, Rosenecker J. Methodological
optimization of polyethylenimine (PEI)-based gene delivery to the lungs
of mice via aerosol application. J Gene Med. 2005 January; 7(1):59-66.;
Rudolph C, Schillinger U, Ortiz A, Plank C, Golas M M, Sander B, Stark H,
Rosenecker J., Aerosolized nanogram quantities of plasmid DNA mediate
highly efficient gene delivery to mouse airway epithelium., Mol Ther.
2005 September; 12(3):493-501). 24 hours after administration of the
aerosol according to the invention containing magnetic-particles, the
mice (n=3) were sacrificed and the amount of plasmid DNA in the lungs was
determined by means of a real-time PCR. The presence of the permanent
magnets on the thorax led to a 2.5-fold higher deposition of plasmid DNA
in the lung (cf. FIG. 21). These results show that permanent magnets are
also suitable for accumulating medicinal substances in aerosols
containing magnetic particles in the lung in vivo.

Example 5

In Vivo Studies with Anaesthetized Domestic Pigs

[0119]Two anaesthetized domestic pigs (approx. 20 kg) were respirated
under controlled conditions, and during the respiration an aerosol
according to the invention containing magnetic particles as described in
Examples 1, 2 and 3 was administered (10 ml of fluidMAG-PEI, 50 nm, 25
mg/ml). On the first pig, a permanent magnet (4 magnets 8×4×2
cm lying one above the other, 0.8 T) was placed on the right side of the
breast, and no magnet was placed on the second pig during the
administration. After the administration of the aerosol, the pigs were
sacrificed and the lungs removed. The magnetic particles in the ventral
lung regions were quantified by means of magnetorelaxometry. The number
of magnetic particles in the right lung by means of a magnetic field is 5
times higher than without a magnetic field (see FIG. 22). These studies
show that aerosols according to the invention containing magnetic
particles can also be guided into the lung of large test animals in a
targeted manner.